Environmental DNA and visual encounter surveys for amphibian biomonitoring in aquatic environments of the Ecuadorian Amazon.

Background: The development of anthropogenic activities has generated a decline in aquatic fauna populations, and amphibians have been the most affected. The decline of batrachofauna is concerning, as 41% of all species worldwide are endangered. For this reason, rapid, efficient, and non-invasive biodiversity monitoring techniques are needed, and environmental DNA (eDNA) is one such tool that has been sparsely applied in Ecuador. This technique has allowed scientists generates information on species diversity and amphibian community composition from a water sample. This study applied eDNA-based biomonitoring analyses and visual encounter surveys (VES) as inventory techniques to identify the diversity of aquatic amphibians in the Tena River micro-basin (TRMB). Methods: The experimental design was divided into three components: (1) fieldwork: all amphibians were recorded by the VES technique and water samples were collected; (2) laboratory work: DNA isolation from amphibian tissue samples and eDNA-containing filters, amplification, electrophoresis, and sequencing were performed; (3) Data analysis: a local DNA reference database was constructed, and eDNA sequence data were processed for classification, taxonomic assignment, and ecological interpretation. Results: Using both eDNA and VES, we detected 33 amphibian species (13 with eDNA only, five with VES only, and 15 with both methods). These species belonged to six amphibian families: Hylidae being the richest with 14 species (three eDNA, one VES, and 10 with both methods), followed by Strabomantidae with nine species (six eDNA, one VES, and two with both methods). All families were detected with both methods, except for the Aromobatidae, having one single record (Allobates aff. insperatus) by VES. Individually, eDNA detected 28 species and had a detection probability (DP) of 0.42 CI [0.40-0.45], while VES recorded 20 species with a DP of 0.17 CI [0.14-0.20]. Similarly, using VES, Cochranella resplendens was detected for the first time in TRMB, while with eDNA, four mountain frogs Pristimantis acerus, Pristimantis eriphus, Pristimantis mallii, and Pristimantis sp. (INABIO 15591) previously recorded at 1,518 m.a.s.l. at altitudes below 600 m.a.s.l. were detected. Conclusions: Results obtained in this study showed that eDNA-based detection had a greater capacity to detect amphibians in aquatic environments compared to VES. The combination of VES and eDNA improves the sensitivity of species detection and provides more reliable, robust, and detailed information. The latter is essential for developing conservation strategies in the Ecuadorian Amazon.

Hematologic variables of free-living Leptodactylus luctator with and without hemoparasites and thrombidiform mites in southern Brazil.

It has been suggested that anuran amphibian parasites can cause clinical signs in situations of environmental imbalance. In the family Leptodactylidae, information about hematology is scarce, although these are well-known tools for the diagnosis and prognosis in clinical practice and potential bioindicators of environmental stress. The objective of this study is to describe Leptodactylus luctator hematology, to report the occurrence of hemoparasites and thrombidiform mites, and to compare the hematological variables under the presence and absence of these organisms. Ectoparasites and heparinized blood samples from 40 free-living specimens of L. luctator were collected for analysis. Hematologic variables and total plasma protein were compared between groups with and without hemoparasites and intradermal mites. As results, structures compatible with hemogregarines, Lankesterella sp., five morphotypes of Trypanosoma spp., microfilaria, Aegyptianella sp., an unidentified intraleukocytic hemoparasite, and frog erythrocytic virus (FEV) inclusion bodies were identified in the blood samples, besides Hannemania spp. intradermal mites. The hemoparasite occurrence was higher than previously reported in other anuran families and locations. Also, L. luctator has smaller red blood cells (RBCs) and white blood cells (WBCs), and a hyposegmentation of the neutrophil nucleus, when compared to many other amphibians. White blood cell, neutrophil, and monocyte counts were higher in animals parasitized by mites. There was no correlation between the number of parasitized RBCs and hematologic variables. This study provides anuran hematologic information, in addition to indicating a host reaction to infestation by Hannemania spp. mites, besides constituting the first record of the distribution of hemoparasites and intradermal mites in L. luctator of the study region.

Anuran fauna of the Parque Estadual Carlos Botelho - Núcleo Sete Barras, southeastern Brazil: species composition, use of breeding sites, and seasonal patterns of breeding activity / Anurofauna do Parque Estadual Carlos Botelho - Núcleo Sete Barras, sudeste do Brasil: composição de espécies, uso de sítios reprodutivos e padrões sazonais de atividade reprodutiva

Abstract: The goal of this work was to study the species composition, the use of breeding sites, and the seasonal patterns of breeding activity of the anuran fauna from Parque Estadual Carlos Botelho - Núcleo Sete Barras, state of São Paulo, southeastern Brazil. Fieldwork was carried out from September 2005 to October 2007 through two main methods: active visual search inside a 10 ha- permanent plot and aural and visual search in seven previously selected aquatic breeding habitats. Species richness was related to the sampling effort by means of species accumulation curve and through non-parametric estimators. Thirty-three species distributed in 12 families were recorded, from which 69% are endemic to the Atlantic Forest. Trachycephalus mesophaeus represents a new record for the park. Cycloramphus lutzorum is included as Data Deficient in the IUCN list. The species accumulation curve did not stabilize, showing some tendency to rise. The use of breeding sites by 20 species was compared using cluster analysis, which revealed two major groups: the first with five species (two habitat generalists and three stream specialists) and the second composed by the other species (with different reproductive modes associated with flooded environments). The breeding period of most species was associated to the rainy season (October to February), and only Scinax hayii showed continuous breeding activity during the entire period of study.

Resumo: O objetivo deste trabalho foi estudar a composição de espécies, o uso de habitats reprodutivos e os padrões sazonais de atividade reprodutiva da anurofauna do Parque Estadual Carlos Botelho - Núcleo Sete Barras, estado de São Paulo, sudeste do Brasil. O trabalho de campo foi desenvolvido de setembro de 2005 a outubro de 2007 por meio de dois métodos principais: procura visual ativa no interior de uma parcela permanente de 10 ha e busca visual e acústica em sete habitats aquáticos de reprodução previamente selecionados. A riqueza de espécies foi relacionada com o esforço amostral por meio de curvas de acumulação de espécies e estimadores não-paramétricos. Foram registradas 33 espécies distribuídas em 12 famílias, 69% das quais são endêmicas da Mata Atlântica. Trachycephalus mesophaeus representa um novo registro para o parque. Cycloramphus lutzorum está incluída como Deficiente em Dados na lista da IUCN. A curva de acumulação de espécie não se estabilizou, mostrando alguma tendência de crescimento. A similaridade no uso dos ambientes aquáticos por 20 espécies foi avaliada por meio de uma análise de agrupamento, que revelou dois grupos: o primeiro com cinco espécies (duas generalistas de habitat e três especialistas de riachos) e o segundo formado pelas demais espécies (com diferentes modos reprodutivos associados a ambientes aquáticos lênticos). O período reprodutivo da maioria das espécies esteve associado à estação chuvosa (outubro a março), e apenas Scinax hayii mostrou atividade reprodutiva contínua durante todo o período de estudo.

Morphological characterization and taxonomic key of tadpoles (Amphibia: Anura) from the northern region of the Atlantic Forest / Caracterização morfológica e chave taxonômica de girinos (Amphibia: Anura) da região norte da Mata Atlântica

Abstract: Although anuran tadpoles are widely distributed and abundant in tropical aquatic habitats, there is a lack of taxonomic keys for the Atlantic Forest. Herein, we developed a dichotomous key for identifying the tadpoles for all species with known larval phase and already recorded in the Atlantic Forest north of the São Francisco River. We analyzed discrete characteristics of 1,042 tadpoles encompassing 63 species of 28 genera from 32 localities. The user-friendly key includes illustration and pictures, and it is a significant step towards improving our knowledge of tadpoles of the Atlantic Forest.

Resumo: Embora os girinos de anuros sejam amplamente distribuídos e abundantes em habitats aquáticos tropicais, não há chaves taxonômicas suficientes para as espécies da Mata Atlântica. Aqui, desenvolvemos uma chave dicotômica para identificar os girinos de todas as espécies com fase larval conhecida e já registradas nesse bioma, ao norte do rio São Francisco. Nós analisamos características discretas de 1.042 girinos pertencentes a 63 espécies (28 gêneros) de 32 localidades. Esta chave de identificação inclui ilustrações e imagens, sendo um passo significativo para um melhor conhecimento dos girinos da Mata Atlântica.

Insecticidal activity of Leptodactylus knudseni and Phyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi and Aedes aegypti.

BACKGROUND: Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anurans Leptodactylus knudseni and Phyllomedusa vaillantii was performed against the mosquito species Anopheles darlingi and Aedes aegypti. METHODS: The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations (LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. RESULTS: The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingi was 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 for An. darlingi larvae was much lower (0.4 ppm) than for Ae aegypti (2.1 ppm). CONCLUSIONS: The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.

Insecticidal activity of Leptodactylus knudseni and Phyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi and Aedes aegypti

Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anurans Leptodactylus knudseni and Phyllomedusa vaillantii was performed against the mosquito species Anopheles darlingi and Aedes aegypti. The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations(LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingi was 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 for An. darlingi larvae was much lower (0.4 ppm) than for Ae aegypti (2.1 ppm). The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.

Insecticidal activity of Leptodactylus knudseni andPhyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi andAedes aegypti

Background Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anuransLeptodactylus knudseni andPhyllomedusa vaillantii was performed against the mosquito speciesAnopheles darlingi and Aedes aegypti.Methods The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations (LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species.Results The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingiwas 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 forAn. darlingi larvae was much lower (0.4 ppm) than forAe aegypti (2.1 ppm).Conclusions The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.(AU)

Insecticidal activity of Leptodactylus knudseni and Phyllomedusa vaillantii crude skin secretions against the mosquitoes Anopheles darlingi and Aedes aegypti

Mosquitoes are important vectors of several diseases, including malaria and dengue, and control measures are mostly performed using chemical insecticides. Unfortunately, mosquito resistance to commonly applied insecticides is widespread. Therefore, a prospection for new molecules with insecticidal activity based on Amazon biodiversity using the anurans Leptodactylus knudseni and Phyllomedusa vaillantii was performed against the mosquito species Anopheles darlingi and Aedes aegypti. The granular secretion from anuran skin was obtained by manual stimulation, and lethal concentrations(LCs) for larvicidal and adulticidal tests were calculated using concentrations from 1-100 ppm. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The skin secretions from the anuran species tested caused significant mortality within the first 24 hours on adults and larvae, but differed within the mosquito species. The calculated LC50 of L. knudseni skin secretions against An. darlingi was 0.15 and 0.2 ppm for adults and larvae, respectively, but much higher for Ae. aegypti, i.e., 19 and 38 ppm, respectively. Interestingly, the calculated LCs50 of P. vaillantii against both mosquito species in adults were similar, 1.8 and 2.1 ppm, respectively, but the LC50 for An. darlingi larvae was much lower (0.4 ppm) than for Ae aegypti (2.1 ppm). The present experiments indicate that skin secretions from L. knudseni and P. vaillantii contain bioactive molecules with potent insecticide activity. The isolation and characterization of skin secretions components will provide new insights for potential insecticidal molecules.(AU)

Anurofauna da Floresta Estacional Semidecidual da Estação Ecológica dos Caetetus, Sudeste do Brasil / Anuran fauna of the Semi-deciduous Forest of the Estação Ecológica dos Caetetus, Southeastern Brazil

A Floresta Estacional Semidecidual é considerada uma das formações florestais mais ameaçadas e suprimidas pela expansão agrícola do início do século passado, com sua cobertura vegetal reduzida a menos de 8 por cento da original. Estudos sobre taxocenoses de anfíbios anuros nessa formação florestal ainda são escassos, mesmo quando consideramos as poucas áreas de floresta primária que restaram. Apresentamos neste estudo os resultados de um levantamento de espécies de anfíbios anuros da Estação Ecológica dos Caetetus, uma unidade de conservação que abriga importante remanescente de Floresta Estacional Semidecidual na região Centro-oeste do estado de São Paulo, região considerada prioritária para conservação no estado. Foram registradas 34 espécies distribuídas em nove famílias, dentre as quais Hylidae e Leptodactylidae foram as que apresentaram as maiores riquezas, com 14 e oito espécies, respectivamente. A elevada riqueza de espécies da área, principalmente da família Hylidae, provavelmente está associada tanto à grande heterogeneidade ambiental dessa formação florestal como à sua localização em área de transição entre áreas de Cerrado e de Floresta Ombrófila. A alta riqueza de espécies, quando comparada à de outras áreas de mesma formação florestal, bem como a presença de representantes das famílias Centrolenidae, Craugastoridae e Hylodidae, que são dependentes de habitats florestais, atestam o bom estado de conservação e a importância regional desse remanescente florestal.

Semi-deciduous Forest is one of the most threatened and deforested vegetation types of Brazil due to its land conversion to agriculture in the beginning of the 20th Century; its distribution is now reduced to less than 8 percent of its original extent. Studies about anuran taxocenosis associated to these forests are scarce, even when we consider the remaining areas of pristine forest. We present herein results from an amphibian survey at the Estação Ecológica dos Caetetus, a conservation unit with a singular preserved Semi-deciduous Forest from the Middle-West region of the State of São Paulo, located inside the priority areas for conservation on São Paulo State. We provided here data on species richness, composition, geographical distribution, and natural history notes for each taxa found in the area. We recorded 34 anuran species belonging to nine families, of which Hylidae and Leptodactylidae are the two most speciose families, with 14 and eight species respectively. The high species richness in the area, mainly in the family Hylidae, can be explained by both the high environmental heterogeneity of this remnant and its geographic location between Cerrado savannas and Atlantic rain forest areas. Its high species richness when compared with other localities of the same vegetation type, as well as the presence of forest dependent species in the families Centrolenidae, Hylodidae and Craugastoridae, indicate both the good conservation status and the regional importance of this forest remnant.

Uso dehabitats naturais e antrópicos pelos anuros em uma localidade no estado de Rondônia, Brasil (Amphibia: Anura)

This paper presents a list of anurans species from Jaburi farm, Espigão D'Oeste Municipality, with special focus on the habitats used during the reproduction period in primary forest and pasture areas. Thirty one species were recorded for this locality from the following families: Bufonidae (3), Dendrobatidae (2), Hylidae (17), Leptodactylidae (7) and Microhylidae (2). The majority of species (24) occurred into and around the primary forest. Fourteen species were observed in breeding activity in the pasture areas, while 9 species were finded only inside primary forest.

Neste trabalho é apresentado a lista de espécies de anfíbios anuros da Fazenda Jaburi. Município de Espigão D'Oeste, Estado de Rondônia, enfocando os ambientes utilizados durante a atividade reprodutiva em áreas de floresta primária c campos de pastagens. Foram registradas 31 espécies para esta localidade pertencentes as seguintes famílias: Bufonidae (3), Dendrobatidae (2), Hylidae (17), Leptodactylidae (7) e Microhylidae (2). A maioria das espécies (24) ocorreram no interior e na borda de floresta primária. Quatorze espécies foram observadas em atividade reprodutiva nos campos de pastagens, enquanto nove espécies foram encontradas apenas dentro de floresta primária.

Uso dehabitats naturais e antrópicos pelos anuros em uma localidade no estado de Rondônia, Brasil (Amphibia: Anura)

This paper presents a list of anurans species from Jaburi farm, Espigão D'Oeste Municipality, with special focus on the habitats used during the reproduction period in primary forest and pasture areas. Thirty one species were recorded for this locality from the following families: Bufonidae (3), Dendrobatidae (2), Hylidae (17), Leptodactylidae (7) and Microhylidae (2). The majority of species (24) occurred into and around the primary forest. Fourteen species were observed in breeding activity in the pasture areas, while 9 species were finded only inside primary forest.

Neste trabalho é apresentado a lista de espécies de anfíbios anuros da Fazenda Jaburi. Município de Espigão D'Oeste, Estado de Rondônia, enfocando os ambientes utilizados durante a atividade reprodutiva em áreas de floresta primária c campos de pastagens. Foram registradas 31 espécies para esta localidade pertencentes as seguintes famílias: Bufonidae (3), Dendrobatidae (2), Hylidae (17), Leptodactylidae (7) e Microhylidae (2). A maioria das espécies (24) ocorreram no interior e na borda de floresta primária. Quatorze espécies foram observadas em atividade reprodutiva nos campos de pastagens, enquanto nove espécies foram encontradas apenas dentro de floresta primária.